The major goal of this project is to discover molecular mechanisms that regulate the connectivity of the visual cortex during development. Surprisingly little is known about the molecular substrates which underlie the formation of the highly specific sets of connections which both characterize and underlie the normal functioning of the visual system. This knowledge is critical not only for understanding visual system development, but to also enable us to genetically target specific populations of neurons to determine their function in visual processing or to deliver therapies following disease or injury to the nervous system. (1) We shall identify molecules that are important in regulating the initial sets of input and output connections of primary visual cortex. To do this, we will utilize high-density DNA microarrays to discover genes that are expressed in visual cortex at very early stages in development in the mouse. It is likely that the genes that regulate the initial connections will be differentially expressed between cortical areas at these times. We will prepare biotinylated cRNA samples from visual, somatosensory and auditory cortex from embryonic and neonatal mice. Data will be analyzed to find genes that are consistently enriched in visual cortex compared to other sensory neocortical regions. (2) We shall examine the spatial and temporal expression of promising candidate genes using in situ hybridization. We will determine which populations of neurons express the candidate genes by combining in situ hybridization with anatomical tracing experiments and immunohistochemistry. (3) We shall identify molecules that regulate the activity dependent refinement of connections which occurs during the critical period. We will therefore prepare samples from the visual cortex of mice in the mid-critical period for ocular dominance plasticity and from mature mice. We will analyze the data to find genes that show up- or down-regulation during the critical period compared to expression levels seen in neonates and mature animals. (4) We shall assess whether promising candidate genes identified in our third aim are regulated by visually-driven activity, by performing in situ hybridization on tissue from visually-deprived animals and comparing the signal strengths. In the longer term, the function of genes which show a spatial and/or temporal distribution pattern which suggests they play a role in the regulation of connectivity will be assessed using in vitro co-culture techniques and anatomical and physiological analysis of transgenic mouse models.